This is a competitive renewal of NIH grant DK33201-19 entitled "Insulin Receptor Substrates and Insulin Action" which focuses on studying the role of phosphorylation, in particular of the insulin receptor substrates and their partners, in the mechanism of insulin action. Over the past four years we have studied the similarities, differences and potentially complementary roles of various insulin receptor substrates in coupling the insulin receptor tyrosine kinase to down-stream effector systems through the creation and characterization of knockout mice and knockout cell lines; studied the role of the IRS proteins in adipocyte differentiation, glucose transport and control of apoptosis; defined a role for SOCS proteins in insulin resistance; and begun to determine the subcellular compartmentalization and differential partnering of the major IRS family members;. This work has led to the realization that IRS proteins mediate different insulin signaling pathways by providing complementary information, that the exact balance of signaling proteins may affect insulin action both positively and negatively, and that these pathways can be altered in a variety of genetic and nongenetic ways, in insulin resistant states. The specific aims for the next five years are to: 1) Define the role of specific IRS proteins and their downstream partners in control of gene expression by using Affymetrix oligonucleotide microarrays and cells lacking specific IRS proteins or in which insulin signaling has been blocked by specific pathway inhibitors. 2) Determine the role of the subcellular compartmentalization and trafficking, and the interaction of the IRS proteins and their downstream partners in intact cells using immunofluorescence confocal imaging, 3-dimensional real time imaging, and FRET analysis. 3) Define the role of the SOCS proteins as inhibitors of IRS mediated signaling in acquired and genetically programmed insulin resistance in vivo using anti-sense and by creating insulin resistance in mice heterozygous for deletion of SOCS-1 and SOCS-3. 4) Determine if the genetic modifiers of insulin resistance in the IR/IRS-1 double heterozygous knockout mouse are similar to or different from those in acquired insulin resistance. Identify the chromosomal loci and the specific genes in these loci which modify insulin resistance in the double heterozygous mouse leading to marked differences in diabetic phenotype in C57B1 and 129Sv mice.